Quantification of Anticougulants in Equine Feed Using Liquid Chromatography-Tandem Mass Spectrometry

A fast screening and simultaneous determination method of four Coumarin-type and one indandione anticoagulants in equine feed was established using LC-MS/MS coupled with solid phase extraction. The fragmentation pathways were studied according to the characteristic fragment ions under electrospray ionization. Due to the weak acidity of the drugs, samples were first infiltrated and deproteinized with methanol, then enriched by solid-phase extraction (SPE) using a MCX cartridge after extracted with 0.02 mol/L NaOH. Detection was performed on a mass spectrometer by negative electrospray ionization (ESI) in multiple reaction monitoring (MRM) mode in five different time segment except for coumarin with positive mode to achieve simultaneously quantitative analysis of five anticoagulants. The limits of detection are 10 ng/g for dicoumarin and warfarin， 4 ng/g for acenocoumarin and coumarin， 15 ng/g for penidione. Limits of quantitation are as low as 10-40 ng/g and the intra-day and inter-day relative standard deviations (RSD) are less than 8.63% and 10.20%, respectively.